Impact of Sugars and Human Milk Oligosaccharides on Infant Microbiome and Obesity Abstract Hispanic children are at high risk for obesity, a disparity that is already established by 2 years of age. Our prior work found that high dietary sugars in early-life contributes to this increased risk and that this adverse effect was obliterated in children who were exposed to extended (not exclusive) breastfeeding for >12 months. Thus, sugars and extended breastfeeding exert opposing effects on early obesity-risk, but the mechanisms are unclear. One way that dietary sugars and extended breastfeeding could impact infants? obesity-risk is by affecting gut microbiome development, which is rapidly evolving during the first 24 months of life. This is a plausible mechanism given that the gut microbiome is implicated in the development of obesity, and that gut microbial changes have been documented in response to dietary sugars and factors in breast milk. One factor in breast milk that may be directly relevant is the mixture of different human milk oligosaccharides (HMOs), which reach the colon intact and serve as prebiotics, shaping the diversity of the gut microbiota. Therefore, the effects of HMOs on microbiome development could be one mechanism by which breastfeeding protects against obesity in infants, a novel concept supported by our preliminary data. Exposure to high dietary sugars in infancy could also impact obesity risk by disrupting cognitive function and appetite regulation, and evidence suggests that these effects might also be mediated by the gut microbiome, and that HMOs, can protect against these impairments, but human studies are lacking. We propose to examine these concepts in a cohort study in 240 Hispanic women and their newborn infants. Participants will be followed from birth to 24 months, with frequent sampling and assessment of breast milk for HMO composition, maternal and infant microbiota, maternal and infant diet, and infant eating behaviors. The primary outcome will be infant body fat by DEXA and secondary outcomes will be infant cognition and appetite regulation, including gut-derived appetite hormones in a sub-set. We have 4 Specific Aims: 1) Determine the joint effects of dietary sugars and HMOs on development of the infant gut microbiome; 2) Determine the joint effects of dietary sugars and HMOs on development of infant body fat.; 3) Determine the joint effects of dietary sugars and HMOs on infant cognitive outcomes, eating behavior and appetite regulation; and 4) Determine whether the effects of dietary sugars and HMOs on infant outcomes are mediated by changes in the gut microbiome. This study will move the field forward by identifying how early-life dietary exposures (focusing on dietary sugars, breastfeeding and HMOs) affect gut microbiome development, and how this affects development of obesity, cognition and appetite regulation. Findings are expected to have significant implications for identifying specific HMOs and/or gut microbial changes that will be protective for obesity and inform future novel intervention modalities applicable to Hispanic women and their infants.